Project Summary Dengue virus (DENV) is endemic in many regions of the Americas where Zika virus (ZIKV) is emerging. These two viruses are closely related genetically and antigenically, so immunity to DENV may influence the outcome of ZIKV infection. The goal of this project is to understand the impact of pre-existing DENV immunity induced by infection or vaccination on ZIKV infection during pregnancy. This question is important because it has been hypothesized that the unexpectedly high rate of adverse consequences associated with ZIKV infection during pregnancy may be the result of previous infection with one of the four DENV serotypes. Primary infection with one DENV serotype is protective against secondary infection with the same serotype. Secondary infection with a different serotype can lead to enhanced disease because cross-reactive, inadequately neutralizing, an- tibodies can facilitate enhanced viral replication and skewed immune responses. DENV and ZIKV are similar enough in the envelope protein (the major target of these enhancing antibodies) that ZIKV could theoretically function as a ?fifth? DENV serotype. This has been explored to some degree, but controversy remains, as some groups have reported cross-protection while others have reported the potential for enhanced disease. Critically, interactions between DENV and ZIKV have not been explored in the setting of pregnancy, even though congenital ZIKV infection is associated with birth defects. Accordingly, through this NIH/ NIAD R01 we will use our nonhuman primate model of ZIKV infection to evaluate whether the severity of ma- ternal and fetal ZIKV infection during pregnancy are enhanced by previous exposure to DENV. There are two Specific Aims: Specific Aim 1. Define the impact of prior DENV infection on the severity of maternal and neonatal ZIKV infection in pregnant macaques. Specific Aim 2. Define the impact of prior DENV vaccination on the severity of maternal and neonatal ZIKV infection in pregnant macaques. In these studies, we will contrast maternal viremia, immune responses, and fetal outcomes with those in DENV-naive individuals infected identically with ZIKV. Our strategy to induce DENV-specific immune re- sponses includes exposure to both wildtype DENV and a leading DENV vaccine candidate, Sanofi Pasteur?s Dengvaxia. These studies are critically important because 1) ZIKV is circulating in many locations where DENV is hyperendemic and 2) Dengvaxia is currently licensed for use, and licensure of other DENV vaccines is imminent, in areas where ZIKV is circulating. By definition, large-scale vaccination campaigns will increase the prevalence of DENV immunity. Whether immunization with Dengvaxia?which simultaneously exposes an individual to all four DENV serotypes?can lead to more severe ZIKV disease is unknown. The results from these experiments will provide important answers to an epidemiologically relevant question; is it safe to vacci- nate against dengue where ZIKV also is co-endemic?